Human induced pluripotent cells (hiPSC) hole enormous promise for regenerative medicine. We propose to explore new methodologies to generate hiPSC without the use of transgene vectors and to develop ways to understand the kinetics, biochemistry and molecular biology of the reprogramming process. In the first part, we have built systems where individual or combinations of reprogramming factors can be controlled artificially and robustly. This will allow us to address questions regarding the temporal sequence of factor requirement, the duration of factor expression necessary for successful reprogramming and other issues. This system will also set the stage for both broad and focused chemical genomics approaches to identify chemical compound that replace individual or combinations of reprogramming factors. The second part builds of results from the first and will explore the dynamics of reprogramming at the epigenetic, transcriptional, steady state mRNA and proteomic levels. This will provide key mechanistic insights into how reprogramming occurs and into the very nature of the pluripotent state. PUBLIC HEALTH RELEVANCE: The proposed studies will provide a better understanding of the reprogramming process by which adult human cells can be converted to induced pluripotent cells (hiPSC). These cells can be derived from patients and will provide important insights into the etiology of complex diseases and potential cell replacement therapies in the future. Other proposed studies will provide more efficient and safer, clinically applicable approaches to generate hiPSC.